Abstract Osteoarthritis (OA) is a leading cause of disability in the US; but despite its prevalence, there is no optimal treatment for this disease. OA is a whole joint disease characterized by articular cartilage destruction, osteophyte formation as well as synovitis. It is associated with local inflammation, which promotes the expression of cartilage-degrading enzymes such as matrix metalloproteinases (MMPs) and apoptosis mediators such as caspases, leading to matrix degradation and cell death. Recently, certain macrolide antibiotics that are typically used to treat bacterial infection, such as erythromycin and azithromycin, have been reported to have anti-inflammatory functions that are independent of the antibiotic function. However, whether these antibiotics have any effects on joint cartilage protection and OA progression is not known. To efficiently evaluate multiple macrolides across several dosage schemes, a sensitive screening method is necessary to narrow down targets for further detailed histological confirmation. Our long-term goal is to investigate and develop new strategies in treating OA. The goal of this research is to screen macrolide antibiotics for the potential to inhibit joint destruction in OA using the Near Infrared Fluorescence (NIRF) imaging technology. This is based on our recent studies demonstrating a chondroprotective role for erythromycin. In addition, by using an MMP-activatable fluorescent probe, we found NIRF signals exhibit gradual changes over the course of OA development in the murine joint that are not always evident from histological analysis. This demonstrates that NIRF imaging is a sensitive approach to detect changes in the joint in various stages of OA that may otherwise go unnoticed. As the same live animals can be imaged quickly and repeatedly, NIRF imaging is also efficient and allows the trajectory of joint destruction in the same animal to be evaluated. Therefore, we hypothesize that other members of the macrolide class of antibiotics can also regulate OA progression as identified through the sensitive NIRF imaging technology. To test this hypothesis, we will evaluate the effect of macrolide antibiotics on joint degeneration in murine experimental OA in vivo and human articular chondrocytes in vitro using NIRF imaging. In addition to commonly used macrolide antibiotics, we will also investigate a derivative of these macrolides that is devoid of its antibiotic function. Our interdisciplinary team consists of investigators with extensive experience in joint pathology and imaging, synergizing our efforts toward this novel research. If successful, this work will open a new direction to explore OA treatment options, capitalizing on drugs already approved for other indications. It will also further develop the sensitive metabolic imaging tool towards in vivo screening of OA therapeutics, thus augmenting traditional evaluation approaches in a profound and complementary way.